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MEMBRANE TRANSPORTERS, ION CHANNELS, AND PUMPS

Regulation of water permeability in rabbit conjunctival epithelium by anisotonic conditions

¹Department of Ophthalmology and ²Department of Physiology and Biophysics, Mount Sinai School of Medicine, New York, New York

Submitted 26 May 2005 ; accepted in final form 22 November 2005

Effects of unilateral exposure to anisotonic conditions on diffusional water permeability of the isolated rabbit conjunctiva were determined. A segment of the bulbar-palpebral conjunctiva was mounted between Ussing-type hemichambers under short-circuit conditions. Unidirectional water fluxes (J_dw) were measured in either direction by adding ³H₂O to one hemichamber and sampling from the other. Electrical parameters were measured simultaneously. J_dw were determined under control isosmotic conditions and after introduction of either hyper- or hypotonic solutions against the tear or stromal sides of the preparations. In each of these four separate conditions, the anisotonic medium produced an 20–30% reduction in J_dw across the tissue, with the exception that to obtain such reduction with increased tonicity from the stromal side (medium osmolality increased by adding sucrose), conditions presumptively inhibiting regulatory volume increase mechanisms (e.g., pretreatment with amiloride and bumetanide) were also required. All reductions in J_dw elicited by the various anisotonic conditions were reversible on restoration of control tonicity. In experiments in which preparations were pretreated with the protein cross-linking agent glutaraldehyde, anisotonicity-elicited reductions in J_dw were not observed. Such reductions were also not observed in the presence of HgCl₂, implying the involvement of aquaporins. However, it is possible that the mercurial may be toxic to the epithelium, preventing the tonicity response. Nevertheless, from concomitant changes in transepithelial electrical resistance, as well as [¹⁴C]mannitol fluxes, [¹⁴C]butanol fluxes, and Arrhenius plots, arguments are presented that the above effects are best explained as a cell-regulated reduction in membrane water permeability that occurs at the level of water-transporting channels. Presumably both apical and basolateral membranes can downregulate their water permeabilities as part of a protective mechanism to help maintain cell volume.

unidirectional water fluxes; net water fluxes; Ussing chamber; short-circuit current; electrolyte transport; cell volume regulation; paracellular mannitol permeability

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